DT-diaphorase (DTD) is an enzyme that is overexpressed in many types of cancerous tissues, including breast, colon, liver, bladder, stomach, the central nervous system (CNS) and lung tumours and in melanomas. Various quinone pro-drugs have been studied to determine their effect on cancer cells expressing DTD. By way of example, the expression of DTD is increased up to 80-fold in primary non-small cell lung cancer (NSCLC) relative to normal lung and up to 400-fold in NSCLC relative to small cell lung cancer (SCLC) cell lines. DTD is also known to activate quinone based pro-drugs and this has been proposed as an approach for selectively targeting cancer cells that express DTD. However, while quinone based pro-drugs have been designed and tested to try and exploit this biology, they have to date been found to suffer from one or more disadvantages.
By way of example, non-small cell lung cancer xenografts with high DTD activity have been shown to be susceptible to the quinone pro-drug mitomycin C. However, although mitomycin C has been shown to have activity in the treatment of non-small cell lung cancer, it is a comparatively poor substrate for DTD (Beall et al., Cancer Research, 54: 3196-3201, 1994) and the metabolism of mitomycin C by DTD is pH-dependent leading to pH-dependent inhibition of DTD at higher pHs (Siegel et al., Mol. Pharmacol., 44:1128-1134, 1993).
Apaziquone or (E)-5-(1-Azirinyl)-3-(hydroxymethyl)-2-(3-hydroxy-1-propenyl)-1-methyl-1H-indole-4,7-dione is an indolequinone that is related to mitomycin C that is also susceptible to reductive conversion to active metabolites by DTD. It has been the subject of clinical trials for the treatment of superficial bladder cancer. However, while Apaziquone is efficiently reduced by DTD, it primarily generates reactive oxygen radicals rather than DNA crosslinks and has been shown to exclusively form DNA strand breaks. Being a conventional “hypoxic-targeting” agent, Apaziquone was not active in spheroid studies demonstrating increased expression of DTD towards the necrotic centre of the spheroid. The results probably relate to the poor ability of Apaziquone to penetrate cells and cross cellular membranes (Bibby et al., Int. J. Oncol., 3: 661-666, 1993).
MeDZQ (2,5-diaziridinyl-3,6-dimethyl-1,4-benzoquinone) is a further example of a promising agent that utilizes the DTD enzyme-directed approach to cancer treatment. Initial results showed that it is more than 150-fold more effective as a substrate for recombinant human DTD than mitomycin C, that it has pH-independent metabolism and did not inactivate DTD at higher pHs (Beall et al., supra; Ross et al. Oncol. Res., 6: 493-500, 1994). Although MeDZQ undergoes bioreductive activation by DTD, it is distinctly different from Apaziquone, as it only produces a very modest increase in cytotoxicity under hypoxic conditions relative to aerobic conditions. Moreover despite displaying a good correlation between DTD activity and MeDZQ cytotoxicity across a range of human tumour cell lines, the formulation of MeDZQ as a useful therapeutic was hampered by its poor solubility.
U.S. Pat. No. 6,156,744 discloses a water soluble quinone pro-drugs, 2,5-diaziridiny 1-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone, referred to as “RH1” and its esters formed with acetyl, benzoyl, naphthoyl groups and protected amino acids. U.S. Pat. No. 6,156,744 shows that RH1 is reduced by DTD and selectively kills cells expressing DTD by crosslinking DNA, and that it is more soluble than MeDZQ.
It remains a problem in the art in developing pro-drugs that are activated by DTD and have pharmacological properties to make them effective drug candidates.